Characterization of helical junction zones in gelatin networks: A spectroscopic approach

Reversible physical crosslinks (junction zones) in gelatine gels or films are thought to consist of triple helical collagen-like helices. Using circular dichroism (CD) the structural elements present in gelatine have been studied. The recognition and quantification of the most important conformations in the preparations have been investigated using a recently developed Library of reference spectra. The spectra of gelatines at low temperature result from extended helical conformations of the polyProline-II type. The prolyl chromophore was shown to have a pronounced influence on the shape and position of the CD helix spectra. Thus, it was concluded that only polypeptides with the appropriate prolyl content can be used as references, while analysing the relative secondary structure contents by CD. Gelatine samples measured at high temperature result from high amounts of 'random coil'. The commonly used reference spectra for random coils deviate significantly from those of denatured collagen (and other denatured proteins). Denatured gelatine is proposed to be a more general reference for random structures. We described different gelatines using CD approaches to distinguish between extended polyProline-II type single and collagen-like triple helices. Also the issue of cis-trans isomerization will be addressed. Finally the results of fluorescence spectroscopy measurements will be described. Using fluorescence quenching, potentially the extent of crosslinking can be determined. Together with resonance energy transfer the quantification of various helical forms (single, multiple: intra- and intermolecular) can be established. Ultimately this analysis would create possibilities to investigate the relation between conformation and bulk mechanical properties for the testing of gelatines.